Hepatology highlights
Authors: Harvey J. Alter
Journal: Hepatology (2004)

After a long asymptote, interest in hepatitis B virus (HBV) therapy is in a logarithmic growth curve. This ascendancy is predicated on a plethora of new and potent nucleoside and nucleotide analogs that have been licensed or are sitting in the Food and Drug Administration pipeline. van Bömmel et al. report a comparison of two acyclic nucleotide analogues, adefovir (recently licensed) and tenofovir (approved for use against HIV), in 53 lamivudine-resistant patients with more than 6 log10 copies/mL of HBV DNA. In contrast to lamivudine, adefovir and tenofovir rarely lead to resistance. Indeed, in this study, no resistance to tenofovir was observed in 35 patients treated for up to 130 weeks. Both drugs were able to significantly reduce HBV DNA in lamivudine-resistant subjects, but tenofovir did it more rapidly, more consistently, and in a significantly higher proportion of patients; by 44 weeks, all tenofovir-treated patients had undetectable HBV DNA compared with only 44% in the adefovir group. In HbeAg+ patients, the decline at 48 weeks was 5.6 logs for tenofovir versus 2.5 logs for adefovir (P < .0001) (Fig.). In tenofovir-treated versus adefovir-treated subjects, 35% and 19%, respectively, lost HbeAg, and 14% and 6%, respectively, lost HBsAg, although the numbers were small in each comparison. Since adefovir and tenofovir are very similar molecules, the differences may be more apparent than real and the authors suggest that similar efficacy might be achieved if adefovir is given at a higher dose. Remarkably, no significant side effects were noted for either drug, resistance was nil, and efficacy was not affected by coexistent HIV infection. It all sounds too good to be true. So where's the catch? The major catch is that these nucleotide analogs are not curative and need to be given continuously to prevent disease recurrence. The fact that viral suppression can now be achieved for extended periods without the emergence of resistant strains is a major breakthrough, but most feel that eradication of HBV infection will require concomitant or sequential immunotherapy. Investigations are now underway to see if a therapeutic HBV vaccine approach could break tolerance and induce immune clearance once the viral load has been minimized. This is a speculative approach, and one has to have faith in its reasonableness. I, for one, am a B-liever. We will ccc who is correct. (See HEPATOLOGY 2005;41:1421–1425.) Next to world peace and hair restoration for bald writers, nothing is more sought after than a noninvasive way to accurately assess liver fibrosis. Hence, Fibroscan was developed to perform one-dimensional transient elastography or liver stiffness measurement (LSM). In this method, a probe that includes an ultrasonic transducer mounted on a vibrator emits a low frequency vibration directed through the intercostal space. The vibration induces an elastic shear wave that propagates through the liver while pulse-echo ultrasound acquisitions measure its velocity. Velocity is directly related to liver stiffness and, basically, the harder the tissue, the faster the wave propagates. The results are only considered valid if 10 successful acquisitions are obtained and the median of these 10 results represent the liver elastic modulus measured in kilopascals (kPa). An exam can be completed in 5 minutes. Appropriate cutoffs were established by receiver-operator curve (ROC) analysis, and stiffness scores in 251 patients were compared with near simultaneous histologic fibrosis scores. Median liver stiffness showed a progressive and significant increase with each METAVIR fibrosis (F) stage, but error bars showed considerable overlap, except for cirrhosis that was clearly separated. Importantly, in a multivariate analysis, fibrosis was the only parameter that significantly correlated with liver stiffness. Values for sensitivity, specificity, likelihood ratio, and positive (PPV) and negative (NPV) predictive values according to fibrosis stage are shown in the table. NPV for fibrosis of F2 or more was 0.56; for F3 or more, 0.93; and for F4, 0.97. Thus, the test had a 93% chance of excluding severe fibrosis (>F3) and a 97% chance of excluding cirrhosis, but it had a considerably lower exclusionary value for F2. The primary value of a noninvasive fibrosis marker is not to predict cirrhosis but to distinguish persons with F2-F3 who are candidates for treatment from patients with F0-F1 who may not need treatment. Stiffness measurements were far from perfect, but they did reasonably well in this respect. Of the 251 patients, 101 (40%) had an LSM of 8.7 kPa or more, suggesting fibrosis of F2 or more; on biopsy, only 9 of these 101 were F1, and none were F0. Treating these 101 patients without biopsy would have led to minimal error, especially since many would routinely treat patients with F1 fibrosis. Although liver biopsy remains the gold standard, it has many pitfalls, including pain, serious complications, poor patient acceptance, and sampling error. LSM obviates the first three, and, importantly, samples an area 100 times the volume of the liver biopsy. Further, it can be repeated frequently allowing serial determinations of fibrosis progression. Percutaneous liver biopsies may have already attained their peak performance characteristics, whereas LSM is in its infancy and will ultimately sample larger or multiple areas in more sensitive ways. I am encouraged that LSM could spare many patients an invasive procedure while maintaining acceptable diagnostic accuracy. At the very least, LSM will add a whole new dimension to the vibrator industry. (See HEPATOLOGY 2005;41:48–54.) Measurement of T regs has rapidly moved from dinosaur status to the forefront of immunological understanding of viral persistence and autoimmunity. T regs constitute 5% to 10% of CD4+ T cells and constitutively express CD25, the interleukin 2 (IL-2) receptor alpha-chain. The study of Cabrera and colleagues serves as a veritable mini-textbook of immunology and immunological techniques. It is well worth reading both for the immunologically adept and those, like myself, who are immunologically inept (as opposed to immunodeficient). This study confirms and extends another recently highlighted article by Sugimoto et al. (HEPATOLOGY 2003;38:1437–1448). The findings in this study are numerous and can be summarized as follows: (1) A significantly higher proportion of CD4+CD25+ cells were found in the peripheral blood (PB) of patients with chronic hepatitis C virus (HCV) infection than in HCV-recovered patients (mean, 3.02% vs. 1.64%; P = .001). (2) The frequency of T regs was higher in PB than in lymph nodes or in liver parenchyma—this was an unexpected finding and might reflect that the liver tissue was obtained from livers with cirrhosis prior to transplant. (3) Cell surface phenotypes suggest that T regs are an early, antigen-primed, memory T-cell population susceptible to apoptosis. (4) HCV-specific interferon γ (IFN-γ) responses as measured by ELISPOT were enhanced by depletion of CD4+CD25+ cells and inhibited in a dose-dependent manner by enrichment with T regs in both chronic and recovered patients (Fig.). (5) CD4+CD25+ cells suppressed HCV-specific CD8+ T-cell proliferation in a dose-dependent manner as assessed in tetramer assays. (6) CD4+CD25+ T regs secreted transforming growth factor β-1 (TGF-β1) and IL-10 and the suppressive effect of T regs could be neutralized by anti–TGF-β1. (7) Transwell cultures showed that the suppressive effect of T regs required cell-to-cell contact. (8) CD4+CD25+ cells respond specifically to HCV antigens by IL-10 production, a T-helper 2 (Th2) response known to suppress Th1 T-cell helper function. (9) T reg frequency correlated with HCV RNA level but not with alanine aminotransferase (ALT) level or fibrosis scores. Thus, in a systematic series of experiments the authors support their hypothesis that CD4+CD25+ T cells suppress CD4+ and CD8 T-cell responses to HCV and hence promote viral persistence. This action seems dependent on enhanced expression of IL-10 and TGF-β1 that results in suppressed proliferation of HCV-specific T cells and IFN-γ secretion in a dose-dependent manner that requires cell-to-cell contact. A key component may be interactions between TGF-β1 surface expression on T regs and the TGF-β receptor on target CD4 and CD8 cells. Therapeutically, one would like to suppress the suppressors in order to enhance viral clearance, but this runs the risk of causing severe CD8-induced liver damage on the way to cure or inciting autoimmune phenomena. Reestablishing the proper balance between regulation and unfettered action seems the answer. The same solution would apply to most governments. (See HEPATOLOGY 2004;40:1062–1071.) There is increasing evidence that apoptosis accounts for significant cell death in hepatitis C virus (HCV)-infected patients. Apoptosis is mediated by a family of intracellular cysteine proteases called caspases. One of the important substrates of caspase activity is the cytokeratin type 1 family, the cleavage of which contributes to cellular collapse during apoptosis. Cytokeratin-18 (CK-18) is a major filament protein in liver cells. Using a monoclonal antibody (M30) that recognizes a caspase cleavage–generated neoepitope of CK-18, Bantel et al. previously showed caspase activation in 20% of HCV-infected hepatocytes even when these cells showed no morphologic signs of apoptosis and when alanine aminotransferase (ALT) levels were normal. This suggested that caspase activation might be involved in very early liver injury. In this study, the authors examined whether caspase-generated CK-18 fragments could be detected in the serum of HCV-infected patients and whether their extracellular release was associated with liver injury. Using an ELISA assay for measuring the CK-18 fragment in serum, they found only low levels in 7 healthy controls (mean, 173 ± 9 U/L) and high levels (mean, 498 ± 58 U/L) in 59 patients with chronic HCV infection. Although patients with more advanced inflammation and fibrosis had considerably higher serum caspase activity than healthy controls and although this correlated with serum ALT (r = 0.73), overall, serum caspase activity did not distinguish between histological grades or fibrosis stages. However, among HCV-infected patients with normal ALT (27% of the population tested), 56% had elevated levels of caspase-cleaved CK-18, and elevations in this subset significantly associated with more severe fibrosis. The authors imply that this assay might be most useful in patients with normal ALT in which decisions regarding biopsy and treatment are most difficult and that patients with elevated CK-18 cleavage in the face of normal ALT should be monitored more closely and considered for biopsy. This study is small, and some of the data seem contradictory. Nonetheless, since the mechanisms and implications of apoptosis and necrosis are different, it is very important to have markers that distinguish these critical events in hepatocyte death. Clearly, the development of an ELISA that specifically measures apoptotic factors in serum would be a significant advance, and large trials to assess the clinical efficacy of the CK-18 fragment assay are now indicated. These data are encouraging, but this measure of apoptosis is not yet an apotheosis. (See HEPATOLOGY 2004;40:1078-1087.) Yesterday upon the stair I saw a man who wasn't there He wasn't there again today How I wish he'd go away? Yesterday upon the stair I saw a lost virus that appeared to flare The virus was there in macrophages And it appears now, will be around for ages (See HEPATOLOGY 2005;41:106–114.) In a rare prospective study of nosocomial transmission, Forns et al. followed 1301 hepatitis C virus (HCV)-negative patients admitted to a liver unit in Barcelona. Six months later, 6 patients (0.46%) were HCV infected. Phylogenetic analysis and multivariate analysis showed that the primary risk factors were duration of hospitalization of more than 10 days (OR = 35) and having an HCV-positive roommate (OR = 12). The message: Watch whom you room with and, in any case, do not do it for too long. (See HEPATOLOGY 2005;41:115–122.) Women have a lower rate of fibrosis and fibrosis progression than men. Di Martino et al. performed a survey and multivariate analysis of some factors unique to women. Fibrosis progression was significantly higher in nulliparous and postmenopausal women and, in the latter, fibrosis was less in those receiving hormone replacement therapy (HRT). HRT reduced risk to that of the premenopausal state. This finding suggested that estrogen has a beneficial effect on fibrosis—it gives one (meno)pause for thought. (See HEPATOLOGY 2004;40:1426–1433.) Fifteen years after plasma-derived HBV vaccine, anti-HBs titer was less than 10 mIU/mL in 30% of children born to HBeAg+ mothers and 62% of vaccinated children without risk factors. A single boost of recombinant vaccine augmented the anti-HBs response in most but not all. The implications are that booster doses of HBV vaccine may be needed, especially after initial plasma vaccination. However, it is possible that the same boost would have been achieved as an anamnestic response to natural exposure, so the booster issue remains unresolved. (See HEPATOLOGY 2004;40:1415–1420.) In the post–liver transplant setting, the early appearance of HCV-specific CD4 cells by IFN-γ ELISPOT predicted mild recurrent graft hepatitis, as did the presence of IFN-γ–secreting CD8 cells, even in the face of high viral load. The late appearance or absence of these T-cell responses correlated with severe recurrence. It is suggested that cell-mediated immunity studies within the first months after transplantation could predict clinical outcome and identify subjects who might benefit most from antiviral therapy. (See HEPATOLOGY 2005;41:72–81.)